Project Summary Lens, an avascular organ, relies heavily on a network of transporting systems to deliver nutrients and other essential components to bulky lens fibers and excrete wastes. Differentiated nuclear fiber cells never turn over and oxidative stress is a major cause of age related cataracts. Connexin (Cx)-forming gap junction channels play an essential role for the metabolic homeostasis of the lens. Besides gap junctions, connexins form hemichannels, permitting transport of molecules between the cell and its extracellular environment. Connexins are truncated in nuclear fibers and this cleavage is increased with aging and oxidative stress. However, little is known regarding the functional importance and regulation of hemichannels formed by full-length and truncated connexins in lens fibers. Our preliminary studies indicate that hemichannels in cortical lens fibers are activated by mechanical loading, mediate uptake of glucose and glutathione, and exhibit self-protective roles againist oxidative damages. As such, we hypothesize that (1) The hemichannels formed by Cx50/Cx46 are activated by mechanical stimulation in cortical lens fibers and mediate uptake of nutrients/antioxidants, which are delivered to inner cortical and nuclear fibers via gap junctions to maintain cell homeostasis and viability; (2) Functional hemichannels formed by both full length and truncated Cx50/Cx46 exhibit self-protection against oxidative damages. The goal is to understand distinctive, new roles of connexin hemichannels in lens fiber cells under normal physiological and pathological (e.g. oxidative stress) conditions. In this proposal, first, we will determine if connexin hemichannels activated by mechanical loading serve as a major transport pathway facilitating the uptake of nutrients and antioxidants into cortical lens fibers, and the role of integrins in regulating hemichannels. Second, we will test if nutrients/antioxidants uptaken by hemichannels in cortical fibers are delivered through gap junctions to inner cortical and nuclear fibers to meet metabolic needs of cell homeostasis and protect inner fiber cells. Third, we will determine if hemichannels formed by both full-length and truncated Cx50/Cx46 offer a self-protective mechanism against oxidative insult. One of the major innovative aspects is that this proposal aims to uncover a novel role that connexin hemichannels formed by full-length and truncated connexins play in facilitating metabolic function of lens fibers and protecting fiber cells against oxidative damages. We will use established lens primary cultures and retroviral expression in lens in situ, a newly developed dominant negative ex vivo approach, and knockout mouse models. It is our expectation that elucidation of mechanistic roles of connexin channels in lens fibers will provide a better understanding of the general homeostatic process of lens under normal and pathological conditions. The outcomes of our research will be significant because the discoveries should make novel and beneficial contributions to new therapeutic strategies and identify drug targets for the treatment of lens disorders such as age-related cataracts.